Preface Cell is the fundamental unit of life of all living organisms present on the earth. Robert Hook gave the term “Cell” while studying on cork. After discovery of the cell, interest among scientist has developed to understand the structure of the different components present in it and function of the cell. Various chemicals for staining of the cell and its components and techniques were discovered by the Cytologist during seventeenth and nineteenth century to identify chromosomes and to determine the stages of mitosis and meiosis and the life cycles of living organisms. Cytogenetics is science which deals with the study of cell particular chromosome and the function of chromosome in transmitting traits from parents to offspring. Chromosome is the physical carrier of gene which control the development of all living organism. The two key mechanism of cell division i.e. mitosis which is responsible for production of identical cells, and the meiosis is important for reproduction and heredity. Techniques have been developed, upgraded and reformed to examine chromosome of different organisms. Similarly, upgradation in microscope have allowed us to develop better understanding about the structure and function of the chromosome.

Objectives To study working principles of simple microscope Study different parts of simple microscope Introduction This is the first ever microscope. It was founded by Leeuwenhoek in 17th century. He combined a convex lens with a specimen holder, for seeing the magnified images of small objects. It looks like a magnifying glass because it has the capacity to magnify between 200 and 300 times. A simple microscope is a small focal lens used to view the magnified images of small objects. It is just like a hand lens and glass with relatively lower magnification power.

Objectives To study working principles of compound microscope Study different parts of compound microscope Introduction A compound microscope is an optical instrument consisting of two short focal-length convex lenses that are used to observe the highly magnified images of small objects by naked eyes. The compound microscope is capable of magnifying up to 1000 the image of a small object by complex system of lens arrangement. The working distance varies from 0.14 to 4 mm. Compound microscopes generally require the sample to be seen to be sufficiently thin, or translucent, to allow the light to pass. Such microscopes provide high magnification, but the view is two-dimensional.

Objectives Differentiate between fixatives and stains. Describe methods of staining. List the commonly used fixatives and stains and their composition. Describe the uses of above mentioned solutions and stains. Introduction Every time a part of an organism is removed, the action of bacteria will soon disintegrate, dry or putrefy it. They must be kept as close to normal as possible or in other words preserved and fixed in order to study these. Chemicals are required in many practical of Biology. Some of these chemicals preserve and specifically colour certain cell parts while others are used as solutions. You will learn about certain fixatives (preservatives) and stains in this chapter.

Objectives Better understand the process and different stages of mitosis. Prepare your own specimens of onion root in which you can visualize all of the stages of mitosis. Principle The genetic information of plants, animals and other eukaryotic organisms resides in several (or many) individual DNA molecules, or chromosomes. For example, each human cell possesses 46 chromosomes, while each cell of an onion possesses 8 chromosomes. All cells must replicate their DNA when dividing. DNA replication in eukaryotes is followed by the process called mitosis which assures that each daughter cell receives one copy of each of the replicated chromosomes. During the process of mitosis, the chromosomes pass through several stages known as prophase, metaphase, anaphase and telophase. The actual division of the cytoplasm is called cytokinesis and occurs during telophase. During each of the preceding stages, particular events occur that contribute to the orderly distribution of the replicated chromosomes prior to cytokinesis. An onion root tip is a rapidly growing part of the onion and thus many cells will be in different stages of mitosis. The onion root tips can be prepared and squashed in a way that allows them to be flattened on a microscopic slide, so that the chromosomes of individual cells can be observed easily. The super coiled chromosomes during different stages of mitosis present in the onion root tip cells can be visualized by treating with DNA specific stains, like Feulgen stain and Acetocarmine stain.

Objectives Better understand the process and different stages of meiosis. Prepare your own specimens of onion root in which you can visualize all of the stages of meiosis. Principle Meiosis is a type of cell division in which the number of chromosomes is halved (from diploid to haploid) in the daughter cells, i.e., the gametes. It occurs in cell that produce gametes in sexually reproducing organisms. The division is completed in two phases, meiosis I and meiosis II. Meiosis I is a reductional division in which the chromosomes of homologous pairs separate from each other. Meiosis II maintain same number of chromosomes, hence, is called as equational division. Stages of meiosis can be observed in a cytological preparation of the pollen mother cells of the anthers of f lower buds.

Objectives Find out the number and types of gametes and gametic recombination in monohybrid, dihybrid and trihybrid crosses. Principle Gametes are reproductive cells that join together to form a new cell called a zygote during sexual reproduction. Pollen in seed-bearing plants, is the gametophyte producing male gamete. Female gametes (ovules) are included in the ovary of plant. At maturity a sexually reproducing individuals forms the gametes. Homozygote generate only one form of gamete. Homozygous referring to a diploid cell or organism possessing two identical alleles of a specific gene (e.g. AA or aa). Heterozygote generate different types of gametes. Heterozygous referring to a diploid cell or organism possessing two different alleles of a specific gene (e.g. Aa).

Objectives Better understand the monohybrid cross. Calculation of monohybrid phenotypic, genotypic and testcross ratios. Principle Monohybrid cross deals with the study of inheritance of one pair of contrasting characters. The alleles of contrasting character selected in the monohybrid cross will show exact dominance and recessive relationship. It is also defined as a cross is made between two true-breeding parents differing for a single trait, producing an F1 generation. These plants are intercrossed to produce an F2 generation. Monohybrid crosses study led to the Mendel’s law of segregation. Monohybrid cross was first produced in pea by Mendel to explain the “law of segregation”. When parents with distinct phenotypes are crossed, in F1 generation, only one type of plants appears phenotypically. However, segregation can be observed in F2 progeny and the population can be divided into two phenotypes classes. After counting the individuals. It is observed that 75% of them express the dominant phenotype and 25% of them express recessive phenotype.

Objectives Better understand the dihybrid cross. Calculation of dihybrid phenotypic, genotypic and testcross ratios. Principle A cross is made between two true-breeding parents differing in two pairs of genes, producing an F1 generation. These plants are inter-crossed to produce an F2 generation. Mendel second law i.e., Law of independent assortment can be explained by studying the inheritance of two characters at a time, simultaneously. When two phenotypically distinct parents differing with respect to two characters governed by two pair of genes are crossed together. Each pair of genes assorts independently into all possible combination forming four kinds of progeny, two of these are parental type and two are recombinant types.

Objectives Better understand the law of independent assortment. History The law was proposed by Gregor Johann Mendel with the other laws together called as ‘Mendelian laws of inheritance’, in 1866. He used pea plant as a model organism, and studied 7 characters. Principle The Law of Independent Assortment, also known as “Inheritance Law”, states that alleles of different genes assort independently of one another during gamete formation. Independent assortment for two genes can be explained by assuming that the two genes are located in two different chromosomes. The two alleles of a gene will be located in the two homologues of the concerned chromosome. Independent separation of these two pairs of chromosomes at anaphase I of meiosis will lead to the independent segregation of the genes located in them. Thus any allele of one gene is equally likely to combine with any allele of the other gene and pass into the same gamete. Independent segregation of two genes produces four different types of gametes in equal proportion. A random union among these gametes gives rise to sixteen possible zygotes. These zygotes yield a 9:3:3:1 phenotypic ratio, which is known as the typical dihybrid ratio. When two pairs of independent alleles enter into F1 combination, both of them have their independent dominant effect. These alleles segregate when gametes are formed but the assortment occurs independently at random and quite freely.

Objectives Chi-square test for goodness of fit of genetic ratios. Introduction Karl Pearson developed the chi square test of statistical significance which is commonly used in Mendelian and population genetics. Chi square (χ2) test is a test of statistical significance which is used to test the significance difference between observed and expected frequencies or ratios. The purpose of χ2 test is to decide if a set of observed data is according to an expected ratio or if it agrees well with an expected or theoretical distribution. The chi square test of independence allow us to evaluate whether the segregation of alleles at one locus is independent of the segregation of alleles at another locus. To distinguish between inheritance patterns that obey Mendel’s laws versus those that do not, a conventional strategy is to make crosses and then quantitatively analyze the offspring’s. Based on the observed outcome, an experimenter may make a tentative hypothesis. For example, it may seem that the data are obeying Mendel’s laws. Hypothesis testing provides an objective, statistical method to evaluate whether the observed data really agree with the hypothesis. The underlying principles behind a statistical approach is to evaluate the goodness of fit between the observed data and the data that are predicted from a hypothesis. This is sometimes called a null hypothesis because it assumes there is no real difference between the observed and expected values. A high deviation would be found between the observed and expected values. If this occurs, the hypothesis is rejected. Hopefully, the experimenter can subsequently propose an alternative hypothesis that has a better fit with the data. The null hypothesis is a particular claim concerning how the data is distributed. More will be said about the construction of the null hypothesis later. The null and alternative hypotheses for each chi square test can be stated as

Objectives Detection of linkage using Chi-square test for goodness of fit. Introduction The tendency of two or more genes to stay together during inheritance is known as linkage and it is the consequence of the concerned genes being located in the same chromosome. Linked genes do not show independent segregation, as a result the ratios obtained in F2 and test cross generation are significantly different from the expected ratios of 9:3:3:1 and 1:1:1:1, respectively. Detection of linkage χ2 using test In genetic experiments, when the material under study is segregating simultaneously for two characters, it may be of interest to know whether the two characters are inherited independently or they are linked. The χ2 test can be used for this purpose. χ2 test is used in detecting linkage in back crosses and inter crosses. First the χ2 is computed based on the theoretical segregating ratios. If χ2 is not significant it may be concluded that the loci segregate independently. If χ2 is significant we may suspect linkage between the loci. The χ2 is then partitioned into components assignable to different factors.

Objectives Study of gene mapping. Introduction All the genes that are linked together form a linkage group. Genes of a linkage group may be presented on a single straight line in the same order in which they are normally present on the concerned chromosome. The number of different linkage groups in a species is, as a rule, equals to its gametic chromosome number (n). The distance between two neighbouring genes is proportional to the frequency of recombination (%) between them depicted on a line drawing. Such a line drawing depicting the linked genes the recombination frequencies between them is known as Linkage map, Genetic map or Chromosome map. Linkage maps provide information on the genes that are linked together and the frequencies of recombination that may be expected between them. The frequency of recombination between two linked genes cannot exceed 50% which is the frequency in case of independent segregation.

Objectives Better understand the phenomenon of multiple alleles. Introduction The word allele is a general term to denote the alternative forms of a gene or contrasting gene pair that denote the alternative form of a gene is called allele. These alleles were previously considered by Bateson as hypothetical partner in Mendelian segregation. Mendel and his followers used the term ‘allele’ or ‘alleleomorph’ to denote the alternative form of the normal gene. It means the genes for tall and dwarf characters of pea plant are alleles. The former is normal or wild allele and the later is mutant allele. A gene can mutate several times producing several alternative expressions. When three or more alleles are found for any particular gene, these are called ‘multiple alleles’. These occupy the same locus in homologous chromosomes. Multiple alleles are defined as genes that are members of the same gene pair and are located on the same locus. All of them control the same character but each of the allele affects that character somewhat differently than the others. In any diploid organism, a particular gene pair is represented by two alleles of the series, one allele on each of the two chromosomes of a homologous pair. This is because an organism has just two chromosomes of each type. These form a homologous pair. However, in the individuals of a population or species, many alternative forms of the same gene may be present. The grouping of all the possible alleles of a gene pair is defined as a system of multiple alleles and their mode of inheritance is called multiple allelism. The multiple allele system can be represented as A1, A2, A3, A4 etc.

Objectives Better understand the various types of gene interaction. Introduction Gene interaction is the phenomenon of two or more genes that affect each other’s expression in different ways in the development of a single character of an organism. It is the modification of normal phenotypic expression of gene due to interaction between alleles or non-alleles. In such situation, two or more than two genes may interact to give rise to a particular phenotype

Objectives Better understand the phenomenon of probability. Introduction The concept of probability is basis to many principles of classical genetics (segregation, independent assortment etc.) The principals of population genetics are based on the considerations of laws of probability. Probability is the likelihood of occurrence of an event. Trial A trial refers to an opportunity provided for an event to occur, e.g. tossing of a coin once constitute one trial as it provides an opportunity for head or tail to occur. A new borne can be either a boy or a girl.

References Rangaswamy, R. 2016. A Textbook of Agricultural Statistics, New Age International Publishers. Singh, B. D. 2014. Fundamental of Genetics, Kalyani Publishers Singh, P. 2016. Elements of Genetics, Kalyani Publishers Strickberger, M. W. 1968. Genetics. 1968. MacMillan, New York.